There’s been a lot of firsts in the world of genome editing happening lately, from its application in human embryonic stem cells, the identification of a smaller more versatile Cas9, to its upgrade to efficiently using homology-directed repair. Now CRISPR-Cas9 is getting its feet wet with epigenome editing thanks to the clever folks in the Gersbach lab at Duke University.
Epigenome editing consists of a targeting system, typically derived from a genome editing system (such as CRISPR-Cas9) with the catalytic component inactivated (as is the case with dCas9) or removed and function conferred by fusion to a novel effector domain. This enables precise epigenomic modification, only limited by the targeting capabilities of your system of choice.
These systems can be used not only to study single genes but also to investigate interesting regulatory elements. Recently, the Bernstein lab showed that a Histone Demethylase (LSD1) can be fused to TALEs to edit the epigenetic status of enhancers at targeted loci in vivo, where it removed enhancer associated chromatin marks (H3K4me2 and H3K27ac) and inactivated related gene expression.
A TALE based system was also used to study histone methylation via G9A or transcriptional activation via p65, where these changes were able replicate the in vivo response to cocaine addiction. The TALE system has seen many developments since the release of a suite of modular epigenetic and transcriptional effectors related to transactivation and histone modification, but now CRISPR-Cas9 joins the party in its deactivated nuclease form: dCas9.
dCas9 has quickly been adapted for transcriptional control by the Liu Lab, Zhang Lab and Church Lab, and now one of the labs that brought us the first optogenetic CRISPR-Cas9 transcription activation system has taken an interest in a different effector domain. The Gersbach lab’s previous system utilized a VP64 transactivation domain (a transcription activating effector) with dCas9, but now they have switched this out for a histone acetyltransferase (HAT) effector domain in order to study histone acetylation.
Here’s what their epigenome editing system can do:
- It utilizes a p300 HAT effector domain to cause some gene activating H3K27ac.
- The targets induced included: MYOD, OCT4, and the mammalian β-globin locus control region (LCR).
- The system also has a key advantage of only requiring a single sgRNA to manipulate enhancers.
- Comparing VP64 to p300, they found that p300 is much more effective at inducing gene expression when editing distal, as opposed to proximal, regulatory elements.
The very modular p300 core domain can also be swapped into other systems, such as CRISPR orthologs that use alternate PAM sequences. The team then showed off the modular of the p300 core domain by swapping out dCas9 and developing Zinc Fingers and TALEs targeted to the same genes and also a gene of interest in past epigenome editing studies utilizing the TET DNA demethylases. Overall, the CRISPR-Cas9 system offers the ability study histone acetylation in a precise and targetted manner.
Gersbach shares that “…you might be able to use this technique for gene therapy to activate genes that have been abnormally silenced or to control the paths that stem cells take toward becoming different types of cells. These are all directions we will be pursuing in the future.”
Set your precision sights on histones with the modular p300 core domain over at Nature Biotechnology, April 2015